Aquaculture Research, 2010, 41, 321 doi:10.1111/j.1365-2109.2010.02479.x Editorial The special issue of Aquaculture Research is comprised of some of the papers presented at the XIII International Symposium on Fish Feeding and Nutrition, which was held in Floriano¤polis, Brazil, from to June 2008 Since its inception in 1984, the International Symposium continue provide excellence in ¢sh nutrition research and the opportunity for communication among researchers New knowledge in ¢sh nutrition research plays an important role in the development of global aquaculture as well as allows for the production of safe and healthy food for human consumption The widespread interest in the subject of ¢sh and crustacean nutrition was marked by the enthusiasm of the 445 participants from 37 countries The scienti¢c programme of the symposium encompassed a workshop on ‘Sustainable aquafeeds for the third millennium’, followed by 296 oral and poster contributions in the following eight sessions: Protein; Nutrient Requirement and Availability; Nutrition and Gene Expression; Nutrition and Health; Environmental Quality and Feeding Strategies; Fish r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd Quality and Food Safety; Feed Ingredients and Feed Processing; and Broodstock and Larvae Seven invited lecturers, and four review papers from these presentations are published in this special issue Selected contributions were submitted for peer review and the resulting manuscripts are published here The participants earned our appreciation, especially those who gave oral presentations, posters and invited papers The planning and organizing of this symposium was a considerable undertaking for which scienti¢c and local organizing committees met with enthusiasm, energy and strong commitment Our special thanks are due to all the anonymous reviewers for the assistance in reviewing all the papers for this proceeding We hope the articles presented in this special issue will contribute towards the advancement of our knowledge in the fascinating ¢eld of ¢sh nutrition De¤bora M Fracalossi, Organizing Committee Chair Santosh P Lall, Scienti¢c Committee Chair 321 Aquaculture Research, 2010, 41, 322^332 doi:10.1111/j.1365-2109.2009.02174.x REVIEW ARTICLE Protein and amino acid nutrition and metabolism in fish: current knowledge and future needs Sadasivam J Kaushik & Iban Seiliez INRA, UMR 1067, Nutrition, Aquaculture & Genomics Unit, 64310 Saint-Pe¤e-sur-Nivelle, France Correspondence: S J Kaushik, INRA, UMR 1067, Nutrition, Aquaculture & Genomics Unit, Saint-Pe¤e-sur-Nivelle, France E-mail: kaushik@st-pee.inra.fr Abstract Optimising the amino acid supply in tune with the requirements and improving protein utilization for body protein growth with limited impacts on the environment in terms of nutrient loads is a generic imperative in all animal production systems With the continued high annual growth rate reported for global aquaculture, our commitments should be to make sure that this growth is indeed re£ected in provision of protein of high biological value for humans The limited availability of ¢sh meal has led to some concerted e¡orts in ¢sh meal replacement, analysing all possible physiological or metabolic consequences The rising costs of plant feedstu¡s make it necessary to strengthen our basic knowledge on amino acid availability and utilization Regulation of muscle protein accretion has great signi¢cance with strong practical implications In ¢sh, despite low muscle protein synthesis rates, the e⁄ciency of protein deposition appears to be high Exploratory studies on amino acid £ux, inter-organ distribution and particularly of muscle protein synthesis, growth and degradation and the underlying mechanisms as a¡ected by dietary factors are warranted Research on speci¢c signalling pathways involved in protein synthesis and degradation have been initiated in order to elucidate the reasons for high dietary protein/ amino acid supply required and their utilization Keywords: proteins, amino acids, ¢sh, nutrition, metabolism Introduction Protein supply from seafood contributes signi¢cantly to human needs in several geographic areas, especially in 322 the developing world as well as in the emerging economies of the world At a global level, about 45% of all ¢sh consumed by humans, totalling about 48 millions tonnes is farm raised (FAO 2007) Aquaculture thus plays a vital role in supplying products known to have a high biological value to humans (Bender & Haizelden 1957) besides providing healthy long-chain w3 polyunsaturated fatty acids (Sargent1997) From a quantitative point of view, e⁄ciency of protein utilization and muscle protein growth are the most crucial issues Although ¢sh are generally considered better converters of dietary protein, compared with terrestrial vertebrates, given the global context of rapid development of aquaculture and the increasing costs and dearth of protein-rich feedstu¡s, there is an impending necessity for improvements in dietary protein utilization, achievable only by optimising dietary supply in tune with the di¡erent physiological needs of organisms This then necessitates a full understanding of the physiological basis for the requirements and e⁄cient exploitation of available sources to meet such needs Protein/energy nutrition of fish: general considerations Critical assessment of protein requirements have already been made by a number of authors over the past two decades (Cowey & Luquet1983; Bowen1987; Cowey 1994, 1995) The general observation is that ¢sh require a higher level of dietary protein than terrestrial farmed vertebrates The general contentions which have very often been put forward with regard to this high protein requirement of farmed ¢sh are as follows: (i) ¢sh have high ß apparent ý protein needs, the basal energy needs of ¢sh are lower than those of r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd Aquaculture Research, 2010, 41, 322^332 Protein and amino acid nutrition and metabolism in ¢sh S J Kaushik and I Seiliez terrestrial animals, due to the aquatic mode of life, poikilothermy and ammoniotelism Based on comparisons of protein e⁄ciency ratios in a number of farmed animals, it becomes clear that ¢sh and terrestrial animals di¡er only in relative protein concentration in the diet required for achieving maximum growth rate and that there were no or little absolute di¡erences in protein requirements (Cowey & Luquet 1983) As already pointed out by Bowen (1987), ¢sh di¡er from terrestrial animals only in the relative protein concentration in the diet required for maximum growth rate and such di¡erences are explained by the lower energy requirement of ¢sh The contribution of proteins/amino acids towards meeting the energy requirements of ¢sh is considered high Much progress has however been achieved through optimising the digestible protein (DP) to digestible energy (DE) ratios by reducing the dietary DP levels with or without concomitant increase in the dietary non-protein DE supply (Cho & Kaushik 1990; Cho & Bureau 2001) A decrease in DP/DE ratios has indeed proven to be extremely e⁄cient in improving protein utilization and decreasing nitrogenous loses in most farmed ¢sh (Kaushik & Cowey 1991; Cho & Bureau 2001) Of the dietary non-protein DE sources, in most species, fats are well utilized both at the digestive tract level and at a post-absorptive level (Sargent,Tocher & Bell 2002) whereas dietary carbohydrates require heat treatment to improve its digestibility and supply of DE (Bergot & Breque1983;Wilson 1994) Increasing the dietary fat levels has indeed been bene¢cial in bringing down the DP/DE ratios having clear bene¢cial e¡ects in terms of nitrogen utilization in most ¢n¢sh (Lee & Putnam 1973; Kaushik & Oliva-Teles 1985; Hillestad & Johnsen 1994; Manuel Vergara, Robaina, Izquierdo & Higuera 1996; Satoh, Alam, Satoh & Kiron 2004) The latitude of action however appears variable depending on the species, some species bene¢ting more from higher dietary non-protein energy than others In all such cases, a major issue however is the increased fat deposition linked with changes in lipogenic enzyme activities (Dias 1999; Regost 2001) Even if digestible carbohydrates are made available, the metabolic utilization of absorbed glucose is limited in most ¢sh (Moon 2001; Panserat & Kaushik 2002) and the net energy supply is reduced (Bureau 1997; Hemre, Mommsen & Krogdahl 2002) although there are differences between species (Furuichi & Yone1982; Panserat, Medale, Blin, Breque, Vachot, Plagnes.Juan, Gomes, Krishnamoorthy & Kaushik 2000; Shiau & Lin 2001; Enes, Panserat, Kaushik & Oliva-Teles 2008 in press) The lack of control of amino acid catabolism as a¡ected by dietary protein levels is indeed considered to be one major reason for the high protein requirements of ¢sh (Cowey & Walton1989) This is somewhat comparable to what is found in the carnivorous cat, where the high protein requirement is considered to be a consequence of the high obligatory nitrogen losses incurred in the conversion of nitrogen from indispensable amino acids (IAA) to dispensable amino acids (DAA) in the liver and to a slow rate of catabolism of IAA (Taylor, Morris, Kass & Rogers 1998) IAA requirements Quantitative data on amino acid requirements for all 10 IAA are available only for a limited number of species (National Research Council 1993; Wilson 2002; Lall & Anderson 2005; Tibaldi & Kaushik 2005) Given the large number of species of farmed ¢n¢sh and shrimp, we should admit that it is indeed di⁄cult to establish the quantitative requirements for all the 10 IAA for each of the species concerned Measured amino acid requirements of di¡erent species, expressed as a proportion of the diet, show also an apparently high degree of variation (Cowey 1994; Mambrini & Kaushik 1995b;Wilson 2002) One major explanation for this apparent variability in IAA requirement data are linked to methodology issues (Cowey 1995): (i) the mode of expression of data (relative to dietary dry matter or dietary DP or DE level, or in absolute terms per unit metabolic mass per day, etc.), (ii) the composition and type of diet used and whether the ¢sh were able to reach their near maximum growth potential, (iii) the criterion used for the estimation of requirement and (iv) the statistical method used for analysing numerical data on dose^response Besides conventional dose^response curves using di¡erent response criteria such as growth, nitrogen utilization, direct or indirect methods of measurement of amino acid oxidation, metabolic responses, new approaches such as single amino acid deletion or reduction (Fournier, Gouillou Coustans, Metailler,Vachot, Guedes,Tulli, Oliva-Teles, Tibaldi & Kaushik 2002; Green & Hardy 2002; Rollin, Mambrini, Abboudi, Larondelle & Kaushik 2003) or diet-dilution techniques (Liebert & Benkendor¡ 2007) have also been attempted in ¢sh with results con¢rming data obtained by conventional methods A close analysis of reliable data available however points towards some degree of homogeneity between r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 322^332 323 Protein and amino acid nutrition and metabolism in ¢sh S J Kaushik and I Seiliez Aquaculture Research, 2010, 41, 322^332 100 Wt Gain, %Max Wt Gain, %Max 100 75 50 25 Lys, %diet 25 Wt Gain, % Max 80 60 10 12 14 Lys, % CP 40 20 0.5 1.0 1.5 Met, %diet 2.0 80 60 40 Figure Analysis of literature data on lysine and methionine requirements of di¡erent species of ¢sh and shrimp 20 –20 di¡erent species Taking lysine and sulphur amino acids as an example, an attempt was made to a meta- analysis of requirement data As the initial sizes and growth rates and experimental conditions vary between studies, a standardized response as the maximum gain in mass in a given study was used For analysing the dose^response, the four-parameter nutrition kinetics analysis (Mercer 1982) was used Based on data from several studies on requirements of di¡erent species for lysine and sulphur amino acids (methionie1cystine) the di¡erent parameters were computed Calculations were made using data on dietary amino acid levels expressed either as percent of the diet or as percentage of crude protein (% CP) The corresponding dose^response curves are presented in Fig Because the main purpose of dietary protein/amino acid supply is for increasing whole body protein accretion, calculation of daily amino acid increment was used for estimating the IAA requirements of carp and trout (Ogino 1980) Since then, this method has been used by a number of authors for getting at least a rough estimate of IAA requirement pro¢le of several species of ¢sh (Kaushik, Breque & Blanc 1991; Mohanty & Kaushik 1991; Ng & Hung 1995; Kaushik 1998; Kim & Lall 2000; Gurure, Atkinson & Moccia 2007) or shrimp (Teshima, Alam, Koshio, Ishikawa & Kanazawa 2002) From these and several other studies, it appears that the ideal protein would be the one that re£ects the whole body IAA pro¢le of the corresponding species The whole body protein bound amino acid pro¢les are however very much similar between di¡erent species and the amino acids depos- 324 100 100 Wt Gain, % Max 50 0 – 20 75 Met, %CP Table Whole body amino acid composition of di¡erent ¢n¢sh and crustaceans (expressed as g16gN À 1)à Amino acid Finfish Ala Arg Asp Cys Glu Gly His Ile Leu Lys Met Phe Pro Ser Thr Trp Tyr Val 6.17 6.16 9.19 0.96 14.29 6.81 2.47 4.29 7.20 7.38 2.75 4.10 4.37 4.15 4.39 1.01 3.02 4.73 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Shrimp 0.82 0.98 0.85 0.26 2.49 1.69 0.63 0.92 0.70 0.89 0.45 0.47 1.13 0.47 0.54 0.29 0.62 0.53 4.86 6.59 8.37 0.78 12.55 5.03 1.85 3.56 6.13 6.42 2.18 3.44 3.13 3.27 3.18 0.90 3.30 3.95 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.56 1.20 0.34 0.11 1.20 1.32 0.17 0.32 0.47 0.51 0.17 0.27 0.39 0.29 0.14 0.19 0.40 0.61 ÃData from a large number of sources including Deshimaru & Shigeno 1972; Wilson & Cowey (1985); Penìa£orida (1989); [65]Mambrini & Kaushik (1995b); [1]Akiyama et al (1997); [54]Kim & Lall (2000); Alam, Teshima, Yaniharto, Ishikawa & Koshio (2002) ited during growth are also similar between di¡erent teleosts as well as crustaceans (Table 1) It has clearly been shown that there is possibly more apparent variability in AA requirement pro¢les than in the whole bodyAA pro¢les of di¡erent teleosts (Akiyama, Oohara & Yamamoto 1997) It is however reassuring that a study by Green and Hardy (2002) con¢rmed that the requirement pro¢le as proposed by National r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 322^332 Aquaculture Research, 2010, 41, 322^332 Protein and amino acid nutrition and metabolism in ¢sh S J Kaushik and I Seiliez From needs to feeds: developing low or non-fish meal diets One of the major issues a¡ecting the aquaculture industry is the availability of protein-rich feedstu¡s Under intensive ¢sh farming conditions, ¢sh meal 200 AA loss (mg /kgBW/d) Research Council (1993) was found to result in the best nitrogen utilization, compared with that of other pro¢les based on whole body protein or from regression analyses The question remains as to whether the ideal protein really re£ects that of the whole body AA pro¢le Few studies have also looked into the potential of some of the DAA and to the ratios between dietary indispensable to dispensable amino acids (IAA/DAA ratio) (Hughes 1985; Mambrini & Kaushik 1994) By feeding rainbow trout with diets containing varying IAA/DAA ratios and using a number of criteria on protein utilization, a ratio of 57:43 was found to be the most suitable (Green, Hardy & Brannon 2002) Similarly, in gilthead seabream, a dietary IAA/DAA ratio of 1.1 was found to be better than a ratio of 0.8 (Gomez-Requeni, Mingarro, Kirchner, CalduchGiner, Medale, Corraze, Panserat, Martin, Houlihan, Kaushik & Perez-Sanchez 2003) Data available today on amino acid requirements not make a clear distinction between needs for the maintenance and growth components The only complete set of data on maintenance requirements for IAA was made available for rainbow trout (Rodehutscord, Becker, Pack & Pfe¡er 1997) Some other studies have estimated the maintenance needs for individual amino acids in ¢sh (Mambrini & Kaushik 1995a) In a comparative study (Fournier et al 2002), the maintenance requirements for arginine was determined in trout, seabass, seabream and turbot Obligatory nitrogen or amino acid losses under protein-free feeding conditions can be assumed to re£ect the minimum physiological needs for IAA (Young & El-Khoury 1995) Measurement of amino acid losses in ¢sh or shrimp under protein-free feeding or fasted conditions can provide valuable information on the obligatory amino acid losses Drawing from whole body amino acid losses when fed a protein-free diet over 28 days (Fournier et al 2002), we could calculate that there are both quantitative and qualitative di¡erences in endogenous losses in amino acids between rainbow trout and turbot (Fig 2), strongly suggesting di¡erences in protein degradation rates and tissues involved Turbot Trout 150 100 50 Figure Pro¢le of whole body amino acid losses (mg kg À BWday À 1) in turbot and in rainbow trout fed a protein-free diet over weeks (unpublished data from the study by Fournier et al 2002) Figure Lysine and sulphur amino acid contents of selected protein sources compared with the requirements for these amino acids by ¢sh and ¢sh oil are the most common feedstu¡s supplying the essential nutrients (amino acids, fatty acids, minerals and trace elements) vital for growth, health, reproduction and physiological well-being of farmed ¢sh.While the marine capture ¢sheries remains constant, the demand for such feedstu¡s derived from capture ¢sheries is on the increase and the costs are rocketing In this context, replacement of ¢sh meal by alternative protein sources remains a major thrust area of research and much has been accomplished in reducing the level of ¢sh meal in all species (Gatlin, Barrows, Brown, Dabrowski, Gaylord, Hardy, Herman, Hu, Krogdahl, Nelson, Overturf, Rust, Sealey, Skonberg, Souza, Stone, Wilson & Wurtele 2007; Lim,Webster & Lee 2008) Fishmeal is unique in that it is not only an excellent source of high quality protein having an ideal IAA pro¢le for ¢sh and shrimp An example of data on lysine and sulphur amino acid contents of di¡erent plant protein sources in comparison with that of the requirements of ¢sh is illustrated in Fig Fish meal is also a good source of essential fatty acids, minerals and trace elements In choosing alternatives to ¢sh meal, it is then necessary to look r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 322^332 325 Protein and amino acid nutrition and metabolism in ¢sh S J Kaushik and I Seiliez Aquaculture Research, 2010, 41, 322^332 at the amino acid pro¢le, but also at other macro and micronutrients In terms of DP supply, compared with ¢shmeal, there are few ingredients which have similar high protein levels, but however with di¡erent amino acid pro¢les It is now clear that not a single ingredient can totally replace ¢sh meal but that one needs to resort to a mixture of ingredients mimicking the amino acid pro¢le of ¢sh meal It is essential when dealing with alternate protein sources to have precise quantitative data on amino acid availability and the biological value It is also imperative that we choose ingredients whose potential antinutritional factors (Tacon 1997; Francis, Makkar & Becker 2001; Kaushik & Hemre 2008) are limited or reduced with respect to the species concerned We have shown in rainbow trout that soyprotein concentrate can totally replace ¢shmeal resulting in equivalent growth and nutrient utilization, provided some additional methionine is supplied (Kaushik, Cravedi, Lalles, Sumpter, Fauconneau & Laroche 1995) Non-¢shmeal diets incorporating a mixture of di¡erent protein sources are well utilized by rainbow trout (Watanabe, Verakunpiriya, Watanabe,Viswanath & Satoh 1998) but much less so by yellowtail (Watanabe, Aoki, Watanabe, Maita, Yamagata & Satoh 2001) In European seabass (Kaushik, Coves, Dutto & Blanc 2004) as well as gilthead seabream (Benedito-Palos, Saera-Vila, CalduchGiner, Kaushik & Perez-Sanchez 2007; De Francesco, Parisi, Perez-Sanchez, Gomez-Requeni, Medale, Kaushik, Mecatti & Poli 2007), there is much potential to reduce the level of ¢sh meal to a signi¢cant level in ¢n¢sh diets Similarly, much progress has also been made to develop non-¢shmeal diets for shrimp (Amaya, Davis & Rouse 2007) There are misgivings on the potential bene¢ts of supplementing diets with free amino acids (Dabrowski & Guderley 2002) Even very early (Nose, Arai, Lee & Hashimoto 1974; Plakas, Tanaka & Deshimaru 1980), di¡erences between postprandial free amino acid levels between ¢sh fed a protein diet or amino acid based diet In rainbow trout, di¡erences in uptake between protein-bound and free amino acids have also been demonstrated with postprandial blood free amino acid peaks appearing later for protein-bound amino acids (Cowey & Walton 1988) They also showed that incorporation of labelled carbon residues into glycogen and lipid from an amino acid diet was greater than from a whole protein diet, whereas incorporation of radioactivity into tissue protein was higher with the latter In both cyprinids (Nose et al 1974; Murai, Akiyama & Nose 1983) and in shrimp (Lim 1993), adjustment of 326 dietary pH improves the utilization of diets with high levels of synthetic amino acids Increasing the number of meals was proposed to improve amino acid utilization (Yamada, Tanaka & Katayama 1981) in carp In order to reduce absorption of free amino acids while digestion of intact protein occurs, coating amino acids with agar for instance has been found to be e⁄cient resulting in improved nitrogen utilization (Cho, Kaushik & Woodward 1992; Fournier et al 2002) These necessary precautions de¢nitely improves utilization of amino acids supplied in the free form even at very high levels There are indeed several reports showing that crystalline amino acids are well utilized both with semi-puri¢ed (Cho et al.1992; Rodehutscord, Mandel, Pack, Jacobs & Pfe¡er 1995; Fournier et al 2002) and practical diets in several species of ¢sh (Kaushik et al 1995, 2004; Williams, Barlow & Rodgers 2001; Yamamoto, Shima & Furuita 2004; De Francesco et al 2007) In plant-protein-based diets, (Espe, Lemme, Petri & El-Mowa¢ 2006) showed that amino acids are utilized as well as protein bound amino acids even at a 10% incorporation level Amino acid utilization Regulation of feed intake by dietary amino acid balance has been little studied.Whether a dietary amino acid de¢ciency or excess leads to increases in voluntary feed intake over a long term has not been analysed in depth A preliminary analysis shows that single amino acid de¢ciencies (arginine, leucine, luysine, methionine) lead to decreased feed intake (de la Higuera 2001) In European seabass, responses appear to di¡er depending on the amino acid, tryptophan de¢ciency exerting the most signi¢cant depression in voluntary feed intake (Tibaldi & Kaushik 2005) Further insight on the consequences of marginal amino acid de¢ciencies linked with dietary DP levels on short or long-term feed intake is needed to optimize dietary amino acid supply and utilization Nutritional regulation of amino acid metabolism has already been dealt with in detail in a number of in-depth reviews (Walton 1985; Cowey & Walton 1989; Dabrowski & Guderley 2002) At the hepatic level, dietary protein levels appear to exert little e¡ect on amino acid catabolism whereas there is a relatively good response of enzymes of amino acid metabolism to the corresponding amino acid intakes The lack of control by dietary protein levels on amino acid r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 322^332 Aquaculture Research, 2010, 41, 322^332 Protein and amino acid nutrition and metabolism in ¢sh S J Kaushik and I Seiliez oxidation in ¢sh contrasts with what is generally seen in mammals and this is considered to be the major reason explaining teleosts’adaptation to high dietary protein levels Somatic growth involves irreversible transformation of dietary substrates and tissue energy stores into tissue and organs There is de¢nitely a good correlation between somatic growth rate and instantaneous protein synthesis rates (Haschemeyer & Smith 1979; Smith 1981; Fauconneau 1985; Carter & Houlihan 2001) Protein synthesis rates di¡er between tissues and the lowest instantaneous fractional protein synthesis rates are seen in the white muscle and the highest values in active tissues such as the liver or the digestive tract (Fauconneau 1985; Fauconneau & Arnal 1985; McMillan & Houlihan 1989b) like in terrestrial vertebrates (Fig 4) The e⁄ciency of deposition of synthesized protein is however high in the muscle of ¢sh (Fauconneau 1985; Peragon, Ortegagarcia, Barroso, de la Higuera & Lupianez 1992; Carter & Houlihan 2001) Nevertheless, while muscle is the largest component of the lean body mass in ¢sh as in most other vertebrates and muscle protein accounts for close to 50% of the body protein, muscle protein synthesis rates represent only about 20% of the whole body protein synthesis (Fig 5) A good correlation between metabolic rate and protein turnover rates appears to exist across di¡erent species (Young 1991) Inclusion of data from rainbow trout ¢ts well to this general scheme (Fig 6), despite the low metabolic rates reported in ¢sh The energetic cost of protein synthesis in ¢sh is considered to be several fold higher than in mammals (Dabrowski & Guderley 2002) and protein oxidation accounts for the most important source of energy in ¢sh (Weber & Haman 1996) Exerting control over this oxidation and knowledge on the quantitative contribution of individual amino acids towards 35 30 25 20 15 10 Ks, %/day Fed Fasted Re-fed Figure Fractional rates of protein synthesis (%/day) in various tissues of rainbow trout (redrawn from McMillan & Houlihan 1989b) (a) Blood Viscera 2% Liver 1% Adipose 3% 11% Gills 3% Head 13% Muscle 55% Fins 1% Skin 11% (b) (c) Other 25% Other 42% Muscle 22% Muscle 50% Liver 14% Liver 1% Dig.Tract Dig.Tract 39% 7% Figure Relative importance of tissue size (a) compared with protein content (b) and protein synthesis rates in different tissues (c) in rainbow trout 800 Mouse Metabolic rate, kJ/kg/d 700 600 500 Chickens 400 Rat 300 200 100 0 10 20 30 40 Protein turnover, g/kg/d 50 Figure Relation between protein turnover and metabolic rates across species (data for terrestrial animals from Young 1991 and for ¢sh from Fauconneau 1985) this metabolic expenditure are areas worth further investigation Fasting followed by re-feeding induces an increase in protein turnover (Smith 1981; McMillan & Houlihan 1989a) The role of insulin, and insulin-like growth factor (IGF-1) as mediators of the anabolic drive is well described in mammals (Millward 1989, 1995) Current knowledge in higher animals shows that this is accomplished by stimulation of the mammalian target of rapamycin (mTOR), a cell signalling pathway involved in the regulation of initiation of mRNA translation (Garami, Zwartkruis, Nobukuni, Joaquin, Roccio, Stocker, Kozma, Hafen, Bos & r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 322^332 327 Protein and amino acid nutrition and metabolism in ¢sh S J Kaushik and I Seiliez Aquaculture Research, 2010, 41, 322^332 Thomas 2003; Tee, Manning, Roux, Cantley & Blenis 2003; Zhang, Cicchetti, Onda, Koon, Asrican, Bajraszewski, Vazquez, Carpenter & Kwiatkowski 2003) In mammals, amino acids as well as insulin are known to act as regulators of this TOR signalling pathway (By¢eld, Murray & Backer 2005; Nobukuni, Joaquin, Roccio, Dann, Kim, Gulati, By¢eld, Backer, Natt, Bos, Zwartkruis & Thomas 2005; Kim, Goraksha-Hicks, Li, Neufeld & Guan 2008; Sancak, Peterson, Shaul, Lindquist, Thoreen, Bar-Peled & Sabatini 2008) Although the mechanisms of regulation are complex and little understood, we have recently been able to show, in rainbow trout, that insulin and amino acids regulate TOR signalling as in mammals (Seiliez, Panserat, Skiba-Cassy, Fricot, Vachot, Kaushik & Tesseraud 2008b), opening new research perspectives on the molecular bases of amino acid utilization in teleosts Given the dynamic status of protein turnover implying continuous protein synthesis and degradation and because muscle protein synthesis rates are low in ¢sh despite the high e⁄ciency of deposited protein, it is essential to get full insight on the protein degradation pathways In ¢sh, we not yet have a clear idea of the relative importance of the three major proteolytic systems operating in vivo (lysosome, Ca21 dependent and ubiquitin^proteasome dependent) The ubiquitin^proteasome route of protein degradation involves two discrete steps: ¢rst, multiple ubiquitin molecules covalently attach to the protein substrate (Ciechanover 1994; Goldberg 1995) and then these tagged proteins are degraded by the proteasome (Kornitzer & Ciechanover 2000), resulting in peptides of 7^9 amino acid residues (Voges, Zwickl & Baumeister 1999) In mammals, the ATP-dependent ubiquitin^proteasome proteolytic pathway is considered to be the major route of protein degradation involved in skeletal muscle loss and is regulated by the nutritional status (Attaix & Taillandier 1998; Lecker, Solomon, Mitch & Goldberg 1999; Jagoe & Goldberg 2001; Lecker, Jagoe, Gilbert, Gomes, Baracos, Bailey, Price, Mitch & Goldberg 2004) In contrast, in rainbow trout (Oncorhynchus mykiss), the activity of the proteasome in muscle does not change during starvation-induced muscle degradation (Martin, Blaney, Bowman & Houlihan 2002) Furthermore, microarray gene expression analysis in atrophying rainbow trout showed that mRNA levels for the subunits of the proteasome were either not a¡ected or downregulated (Salem, Kenney, Rexroad III & Yao 2006), leading to the suggestion that degradation of muscle proteins in trout occurs by a route distinct from the 328 one observed in mammals (Mommsen 2004) But, our own recent data show that, in the muscle of rainbow trout, the polyubiquitination step of the ubiquitin^proteasome route is regulated by feeding similar to what is observed in mammals (Seiliez, Gabillard, Skiba-Cassy, Garcia-Serrana, Gutierrez, Kaushik, Panserat & Tesseraud 2008a) and supports the idea that we have to reconsider the role of this proteolytic route in muscle protein degradation and its nutritional regulation Future research Given the general context of aquaculture and the importance of dietary protein/amino acids in aquafeeds, a number of areas need our attention.We need to strengthen our understanding of the consequences of marginal amino acid imbalances under minimal dietary DP levels capable of maximum growth and physiological well being Given the relatively high contribution of excess amino acids for energy, we need to gain knowledge on the relative contributions or preferential utilization of individual amino acid oxidation to overall metabolic demands Similarly, understanding the role of individual amino acids directly or indirectly through hormonal factors in eliciting the anabolic drive in the regulation of muscle growth is necessary.While we know that signi¢cant ‘protein-sparing’ and reduction in nitrogenous losses are achieved by decreasing the DP/DE ratios, we need to get more insight on the underlying mechanisms especially as a¡ected by dietary factors Comprehensive data should bring forth more similarities than di¡erences between terrestrial animals and aquatic organisms in their nitrogen metabolism and utilization References Akiyama T., Oohara I & Yamamoto T (1997) Comparison of essential amino acid requirements with A/E ratio among ¢sh species (Review paper) Fisheries Science 63, 963^970 Alam M.,Teshima S.I.,Yaniharto D., Ishikawa M & Koshio S (2002) Dietary amino acid pro¢les and growth performance in juvenile kuruma prawn Marsupenaeus japonicus Comparative Biochemistry and Physiology (Part B Biochemistry and Molecular Biology) 133B, 289^297 Amaya E.A., Davis D.A & Rouse D.B (2007) Replacement of ¢sh meal in practical diets for the Paci¢c white shrimp (Litopenaeus vannamei) reared under pond conditions Aquaculture 262, 393^401 r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 322^332 Aquaculture Research, 2010, 41, 322^332 Protein and amino acid nutrition and metabolism in ¢sh S J Kaushik and I Seiliez Attaix D & Taillandier D (1998) The critical role of the ubiquitin^proteasome pathway in muscle wasting in comparison to lysosomal and Ca21-dependent systems In: Advances in Molecular and Cell Biology (ed by E.E Bittar & A.J Rivett), pp 235^266 JAI Press, Stamford, CT, USA Bender A.E & Haizelden S (1957) Biological value of the proteins of a variety of ¢sh meals British Journal of Nutrition 11, 42^43 Benedito-Palos L., Saera-Vila A., Calduch-Giner J.A., Kaushik S & Perez-Sanchez J (2007) Combined replacement of ¢sh meal and oil in practical diets for fast growing juveniles of gilthead sea bream (Sparus aurata L.): networking of systemic and local components of GH/IGF axis Aquaculture 267, 199^212 Bergot F & Breque J (1983) Digestibility of starch by rainbow trout: e¡ects of the physical state of starch and of the intake level Aquaculture 34, 203^212 Bowen H (1987) Dietary protein requirements of ¢shes-A reassessment Canadian Journal of Fisheries & Aquatic Sciences 44, 1995^2001 Bureau D.P (1997) The partitioning of energy from digestible carbohydrates by rainbow trout (Oncorhynchus mykiss) University of Guelph, Guelph, ON, Canada, 170pp By¢eld M.P., Murray J.T & Backer J.M (2005) hVps34 is a nutrient-regulated lipid kinase required for activation of p70 S6 kinase 280, 33076^33082 Carter C.G & Houlihan D.F (2001) Protein synthesis In: Fish Physiology.Vol 20 Nitrogen Excretion (ed by P.A.Wright & P.M Anderson), pp 31^75 Academic Press, New York, NY, USA Cho C.Y & Bureau D.P (2001) A review of diet formulation strategies and feeding systems to reduce excretory and feed wastes in aquaculture Aquactic Research 32, 349^360 Cho C.Y & Kaushik S.J (1990) Nutritional energetics in ¢sh: energy and protein utilization in rainbow trout (Salmo gairdneri).World Review of Nutrition and Dietetics 61,132^172 Cho C.Y., Kaushik S & Woodward B (1992) Dietary arginine requirement of young rainbow trout (Oncorhynchus mykiss) 102, 211^216 Ciechanover A (1994) The ubiquitin^proteasome proteolytic pathway Cell 79,13^21 Cowey C.B (1994) Amino acid requirements of ¢sh ^ a critical appraisal of present values Aquaculture 124, 1^11 Cowey C.B (1995) Protein and amino acid requirements: a critique of methods 11,199^204 Cowey C.B & Luquet P (1983) Physiological basis of protein requirements of ¢shes Critical analysis of allowances In: IV International Symposium on Protein Metabolism and Nutrition Les Colloques INRA, No 16 (ed by M Arnal, R Pion & D Bonin), pp 365^384 Cowey C.B & Walton M.J (1988) Studies on the uptake of (14C) amino acids derived from both dietary (14C) protein and dietary (14C) amino acids by rainbow trout, Salmo gairdneri Richardson Journal of Fish Biology 33, 295^305 Cowey C.B & Walton M.J (1989) Intermediary metabolism In: Fish Nutrition, 2nd edn, (ed by IJEH.) pp 259^329 Academic Press, NewYork, NY, USA Dabrowski K & Guderley H (2002) Intermediary metabolism In: Fish Nutrition (ed by J.E Halver & R.W Hardy),3rd edn, pp 309^365 Elsevier, Amsterdam, the Netherlands De Francesco M., Parisi G., Perez-Sanchez J., Gomez-Requeni P., Medale F., Kaushik S.J., Mecatti M & Poli B.M (2007) E¡ect of high-level ¢sh meal replacement by plant proteins in gilthead sea bream (Sparus aurata) on growth and body/¢llet quality traits Aquaculture Nutrition 13, 361^372 de la Higuera M (2001) E¡ects of nutritional factors and feed characteristics on feed intake In: Food Intake in Fish (ed by D Houlihan, T Boujard & M Jobling), pp 250^268 Iowa State University Press, State Avenue Ames, IA, USA Deshimaru O & Shigeno K (1972) Introduction to the arti¢cial diet for prawn Penaeus japonicus Aquaculture 1, 115^133 Dias J (1999) Lipid Deposition in Rainbow Trout (Oncorhynchus mykiss) and European Sea Bass (Dicentrarchus labrax): Nutritional Control of Hepatic Lipogenesis University of Porto, Portugal & University of Bordeaux I, France, 190pp., annexes Enes P., Panserat S., Kaushik S & Oliva-Teles A (2008) Nutritional regulation of hepatic glucose metabolism in ¢sh Fish Physiol Biochemistry, in press Espe M., Lemme A., Petri A & El-Mowa¢ A (2006) Can Atlantic salmon (Salmo salar) grow on diets devoid of ¢sh meal? Aquaculture 255, 255^262 FAO (2007) The State of World Fisheries and Aquaculture 2006 FAO, Rome, p.162 Fauconneau B (1985) Protein synthesis and protein deposition in ¢sh In: Nutrition and Feeding in Fish (ed by C.B Cowey, A.M Mackie & J.G Bell), pp 17^45 Academic Press, London, UK Fauconneau B & Arnal M (1985) In vivo protein synthesis in di¡erent tissues and the whole body of rainbow trout (Salmo gairdnerii R.) In£uence of environmental temperature A 82,179^187 Fournier V., Gouillou Coustans M.F., Metailler R., Vachot C., Guedes M.J., Tulli F., Oliva-Teles A., Tibaldi E & Kaushik S.J (2002) Protein and arginine requirements for maintenance and nitrogen gain in four teleosts BritishJournal of Nutrition 87, 459^469 Francis G., Makkar H & Becker K (2001) Antinutritional factors present in plant-derived alternate ¢sh feed ingredients and their e¡ects in ¢sh Aquaculture 199, 197^227 Furuichi M & YoneY (1982) Changes in activities of hepatic enzymes related to carbohydrate metabolism of ¢shes in glucose and insulin-glucose tolerance tests 48, 463^466 Garami A., Zwartkruis F.J., Nobukuni T., Joaquin M., Roccio M., Stocker H., Kozma S.C., Hafen E., Bos J.L & Thomas G (2003) Insulin activation of Rheb, a mediator of mTOR/ S6K/4E-BP signaling, is inhibited by TSC1 and 11, 1457^1466 r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 322^332 329 Aquaculture Research, 2010, 41, 433^449 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al plicatilis, were added twice a day adjusted to 5^10 rot mL À 1in the intensive system and in the semi-intensive system adjusted to 1^2 rot mL À in trial A and to 4^5 rot mL À in trials B and C Rotifers’ feeding period lasted from 2^20 dah in trial A and it was extended until 30 dah in trials B and C Rotifers were growing out on baker’s yeast, Saccaromices cerevisiae and enriched with DHA Protein Selco (Inve Aquaculture, Dendermonde, Belgium) following the manufacturer’ instructions In trials A and B, the weaning protocol included manual feeding from 20 dah (Genma Micro, Skretting, France) four times a day for days and auto- Figure Red porgy larvae excreting live Artemia matic feeding every hour from day 25 In trial C, two co-feeding and early weaning protocols using Artemia and microdiets for di¡erent periods were compared as shown in Table Therefore, the quantity and onset of Artemia Instar I (25^250 A0 L À 1) (AF type, INVE Aquaculture), Artemia Instar II (EG tipe, INVE Aquaculture) enriched with A1 Easy Selco (INVE Aquaculture) and commercial diet onset (Genma Micro) varied according to the co-feeding and early weaning protocols (Table 1) Samples and measurements Larval growth was assessed measuring the total length of 25 larvae every 5^7 days, using a pro¢le projector (Nikon V-12A, NIKON, Tokyo, Japan) Lengthspeci¢c growth rate (SGR) was calculated using the following equation: SGR ([Ln(Lt) À Ln(L0)]/t)  100; where Lt is the larval length at the end of time period t, L0 is the length at the beginning of time period t and t is the time period in days Survival was determined after 95 dah by individual counting of all remaining ¢sh using a fry counter (TPS Fish counter, Type Micro; Impex Agency, Hoerning, Denmark) From 15 dah, dead ¢sh were recorded daily and from that day, survival was accordingly estimated taking into account daily mortality and ¢nal alive ¢sh Table Summary of the rearing parameters and modi¢cations applied in the di¡erent trials Trial Cw Oxygen (ppm) Temperature ( 1C) Photoperiod 24 h: (Natural1artificial) 12:12 h (Natural1artificial) Phytoplankton (age)à SMIS and IS (cells mL À 1) Rotifers (age)à SMIS (ind mL À 1) IS (ind mL À 1) Instar I (age)à SMIS (ind L À 1) IS (ind L À 1) Instar II (age)à SMIS (ind L À 1) IS (ind L À 1) Weaning (onset)à Trial A Trial B W1 W2 6.5 Æ 0.5 20.0 Æ 0.5 6.6 Æ 0.6 20.0 Æ 0.5 6.0 Æ 0.8 20.4 Æ 0.7 6.0 Æ 0.8 20.4 Æ 0.5 2–40 dah 40–50 dah 2–25 dah 250  103 2–20 dah 1–2 5–10 13–20 dah 25–250 25–250 18–50 dah 500 1000 20 dah 10–15% biomass day–1 2–20 dah 20–50 dah 2–25 dah 250  103 2–30 dah 4–5 5–10 13–16 dah 25–250 25–250 15–50 dah 500 1000 20 dah 2–20 dah 20–50 dah 2–25 dah 250  103 2–30 dah 4–5 5–10 13–16 dah 25–250 25–250 15–50 dah 500 1000 20 dah 2–20 dah 20–50 dah 2–25 dah 250  103 2–30 dah 4–5 5–10 13–16 dah 25–250 25–250 15–40 dah 250 500 15 dah ÃValues express the range of larval age in days after hatching wW, weaning protocol; SMIS, Semi-intensive system; IS, Intensive system r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 435 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al Activity test In trial C, an activity test was performed on days 20 (10 s air exposure) and 30 dah (60 s air exposure) Larvae (n 25) in triplicate from each replicate tank were air exposed in a 500 mm nylon mesh screen Later, larvae were transferred to a net container (1L) in their original rearing tank and survival was registered 24 h after the test Biochemical analysis For biochemical analysis, 1000 larvae per tank were collected at 12 and 20 dah and 200 juveniles at 35 and 50 dah from each tank and treatment In order to determine prey nutritional quality, samples of enrichment products, enriched rotifers, enriched Artemia and microdiets were collected and analysed throughout the di¡erent trials (Table 2) In addition, in trial B, samples of the remaining rotifers in the larval tanks were collected before the ¢rst food addition in the morning from both rearing systems on 8, 10 and 12 dah (Table 3) All the biochemical analyses were conducted in triplicate Protein and ash were determined according to AOAC (1990) Total lipids were extracted as described by Folch, Lees and Sloane-Stanley (1957) The FA methyl esters were obtained by transesteri¢cation with H2SO4 (Christie1989) and puri¢ed by adsorption chromatography on NH2 Sep-Pack cartridges (Waters, S.A., Milford, MA, USA) as described by Fox (1990), and separated and quanti¢ed by Gas^ Liquid chromatography as described by Izquierdo,Watanabe,Takeuchi, Arakawa and Kitajima (1990) Aquaculture Research, 2010, 41, 433^449 Table Proximate composition [lipid, protein and ash content; g kg À 1dry weight (dw)] and fatty acid composition (% total fatty acids) of enriched rotifers, Artemia Instar II and microdiets fed to red porgy larvae Live preys Enrich Rotifers % Lipids (dw) % Protein (dw) % Ash (dw) 16:0 16:1n-7 18:0 18:1n-9 18:1n-7 18:2n-6 18:3n-3 20:1n-9 ARA (20:4n-6) EPA (20:5n-3) DHA (22:6n-3) 22.35 52.27 1.20 14.60 11.51 5.46 19.12 3.09 7.77 1.35 2.19 1.83 6.63 10.11 SSaturated SMonounsaturated Sn-3 Sn-6 Sn-9 Sn-3HUFA 23.93 Æ 0.63 24.14 Æ 3.60 39.26 Æ 1.36a 32.89 Æ 1.73b EPA/ARA DHA/EPA DHA/ARA Oleico/DHA Oleico/n-3HUFA n-3/n-6 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Enrich Artemia 2.30a 4.34a 0.27a 0.96a 1.20a 0.56 0.55a 0.17a 1.30a 0.39a 0.10a 0.23a 0.34 0.32 25.45 55.38 0.72 16.19 3.47 6.47 19.06 5.84 5.47 11.31 1.50 1.62 7.56 9.11 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Microdiet 2.15b 2.25ab 0.26b 2.66ab 0.31b 1.65 0.82a 0.25b 0.37a 1.47b 0.09a 0.16a 0.82 1.82 17.09 57.66 17.09 17.48 3.72 4.76 12.23 2.79 21.64 3.51 3.56 0.55 5.98 8.61 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.83c 1.09b 0.83c 1.30b 0.79b 0.71 0.56b 0.85a 3.95b 0.68c 0.67b 0.15b 1.50 1.16 27.19 Æ 1.58 27.59 Æ 1.08c 21.01 10.95 22.78 18.55 Æ Æ Æ Æ 0.52a 31.40 Æ 3.14b 1.23a 9.06 Æ 0.48a 0.68a 21.41 Æ 0.59ab 0.56 18.36 Æ 2.85 20.68 22.83 20.01 15.78 Æ Æ Æ Æ 2.89a 3.57b 1.06b 2.92 3.68 1.53 5.61 1.89 1.03 1.94 Æ Æ Æ Æ Æ Æ 0.59a 0.05a 0.85a 0.07a 0.04 0.21a 11.04 1.48 16.29 1.44 0.80 0.94 Æ Æ Æ Æ Æ Æ 1.21b 0.23ab 3.02b 0.24ab 0.18 0.27c 4.67 1.20 5.59 2.15 1.05 3.46 Æ Æ Æ Æ Æ Æ 0.05a 0.12b 0.60a 0.37b 0.13 0.26b Values (mean Æ SD) followed by di¡erent superscript letters within a row for the same trial were signi¢cantly di¡erent (Po0.05) DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid; ARA, arachydonic acid; HUFA, highly unsaturated fatty acid Statistical analysis All the data were statistically treated using SPSS Statistical Software System ver 15.0 (SPSS 2000, Chicago, IL, USA) A T-test for a simple mean comparison analysis (Po0.05) (Sokal & Rolf 1995) was applied between systems Variances were tested for normality and homogeneity Results are presented as mean standard deviation (Table 4) Results Growth and survival Trial A In trial A, although the average total length of red porgy larvae at 5,10,15,35 and 40 dah was not signif- 436 icantly di¡erent between rearing systems, at 50 dah, the total length of the larvae from semi-intensive system was signi¢cantly higher than that of the intensive system (Fig 2) During the whole rearing period, semi-intensive system larvae had a higher SGR, although without statistically signi¢cant di¡erences (P40.05) between rearing systems Three di¡erent growth periods in terms of SGR were observed: an early rapid growth period (5^15 dah) (5.8^6.7% SGR in the intensive and semi-intensive systems respectively) when prey sources were restricted to rotifers, a second period with a strong reduction in SGR (15 to 30 dah) when new prey types were incorporated (Artemia and microdiets) and a third growth period with increased r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al Unconsumed preys Semi-intensive a Intensive b % Lipids (dw) % Protein (dw) % Ash (dw) 12.37 Æ 1.22 64.41 Æ 13.70 0.91 Æ 0.28a 16:0 16:1n-7 18:0 18:1n-9 18:1n-7 18:2n-6 18:3n-3 20:1n-9 ARA (20:4n-6) EPA (20:5n-3) DHA (22:6n-3) 18.66 9.58 4.02 12.07 2.53 10.26 11.16 1.99 0.83 3.81 4.40 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.33 0.32 0.11 0.47 0.00 0.57 0.94 0.01 0.00 0.10 0.19a 17.60 8.80 4.26 11.91 2.82 10.76 9.99 1.91 0.91 4.10 6.70 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 1.12 0.32 0.41 0.50 0.03 0.78 0.83 0.01 0.04 0.25 0.61b SSaturated SMono-unsaturated Sn-3 Sn-6 Sn-9 Sn-3HUFA 26.68 29.53 24.86 12.35 16.83 11.14 Æ Æ Æ Æ Æ Æ 0.41 1.55 0.78 0.43 0.69 0.15a 25.04 29.63 25.77 13.70 17.17 12.95 Æ Æ Æ Æ Æ Æ 1.65 0.79 1.87 1.24 0.88 0.98b 4.58 1.15 5.28 2.74 1.08 2.01 Æ Æ Æ Æ Æ Æ 0.15 0.02a 0.25a 0.01a 0.03 0.01a 4.51 1.63 7.37 1.79 0.93 1.88 Æ Æ Æ Æ Æ Æ 0.07 0.05b 0.33b 0.24b 0.11 0.03b EPA/ARA DHA/EPA DHA/ARA Oleic/DHA Oleic/n-3HUFA n-3/n-6 16.97 Æ 1.81 71.22 Æ 4.13 0.35 Æ 0.12b Values express average of pool samples from each tank replicate at days 8, 10 and 12 Values (mean Æ SD) followed by di¡erent superscript letters within a row were signi¢cantly di¡erent (Po0.05) DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid; ARA, arachydonic acid; HUFA, highly unsaturated fatty acid Table Growth constants 30 * SMIS-TA IS-TA 25 Total length (mm) Table Proximate composition [lipid, protein and ash content; g kg À dry weight (dw)] and fatty acid composition (% total fatty acids) of unconsumed rotifers in the rearing tank in trial B 20 15 10 0 10 20 30 40 Age (dah) 50 60 Figure Total length evolution (TL) of red porgy (Pagrus pagrus) larvae culture under di¡erent rearing systems in Trial A ÃSigni¢cant di¡erences Larval growth followed an exponential equation TL 2.4677e0.0451d; r2 50.9939 in the semi-intensive system and TL 2.7639e0.0388d; r2 50.9903 in the intensive system, where TL is the total length (mm) and d is the age in days 10 SMIS-TA IS-TA SGR Aquaculture Research, 2010, 41, 433^449 5-15 15-30 30-50 Total (5-50) Age (range) Figure Speci¢c growth rate evolution according to the rearing system in trial A Constants Treatment a b r2 Trial B ISW1-TC SMISW1-TC ISW2-TC SMISW2-TC 3.0394 3.6137 3.2482 3.3708 0.0425 0.0399 0.0404 0.0413 0.9977 0.9967 0.9942 0.9966 From 15 to 50 dah, larvae from a semi-intensive system (Fig 5) were signi¢cantly larger in total length than the intensive system larvae Moreover, these larvae were larger than those obtained in trial A for the same age and rearing system (23.5 Æ 2.7 vs 29.5 Æ 3.0 mm total length) in semi-intensive system 50 old larvae in trial A and B respectively Again, three di¡erent growth periods were found in terms of SGR (Fig 6); however, a signi¢cant improvement in larval growth was obtained in the second period (15^30 dah), in comparison with the previous trial for both rearing systems SMIS, Semi-intensive system; IS, Intensive system; TC, Trial C SGR values (4.5^5.3%) from 30 to 50 dah when larvae were actively fed on a microdiet (Fig.3) Survival up to 50 dah was not signi¢cantly di¡erent between rearing systems 4.4^4.9% in the semi-intensive and intensive systems, respectively, the highest larval mortality period being from to 15 dah (Fig 4) r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 437 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al 100 10 SMIS-TA IS-TA SMIS-TB 60 * 40 20 Hatching 15 20 Age (dah) 50 35 5-15 Figure Survival evolution according to the rearing system in trial A 35 100 * SMIS-TB IS-TB * 20 * 10 * * SMIS-TB IS-TB 80 25 15 15-30 30-50 Total (5-50) Age (range) Figure Speci¢c growth rate evolution according to the rearing system in trial B % Survival Total length (mm) IS-TB SGR % Survival 80 30 Aquaculture Research, 2010, 41, 433^449 * 60 * * 40 * * 35 50 20 0 10 20 30 40 Age (dah) 50 60 Figure Total length evolution (TL) of red porgy (Pagrus pagrus) larvae culture under di¡erent rearing systems in trial B Ãsigni¢cant di¡erences Larval growth follows an exponential equation TL 3.5464e0.0392d; r2 50.959 in the semi-intensive system and TL 3.0267e0.0423d; r2 50.999 in the intensive system, where TL is the total length (mm) and d is the age (days after hatching) Hatching 15 20 Age (dah) Figure Survival evolution according to the rearing system in trial B 35 dah, mesocosm-reared larvae showed an average prey availability that was six times higher (Fig 8) Trial C Overall, survival in the semi-intensive system in trial B was higher than in trial A (4.4% vs 21.8 in trials A and B respectively) Besides, the highest larval mortality period from to 15 dah was noticeably reduced in the semi-intensive system, to reach a 38% higher survival than in trial A for the same age (Fig 7) However, no noticeable improvements were obtained in ¢nal survival for larvae reared in the intensive system (4.9% vs 5.3% in trials A and B respectively) In this trial, the estimation of available prey per larvae in relation to estimated larval density, at di¡erent ages, was re£ected in important di¡erences among rearing systems; thus, for the period from to 438 In trial C, the mean total lengths for 50-day-old larvae were not signi¢cantly di¡erent either between rearing systems or between weaning protocols applied for each system The average total length (25.04 mm) obtained for this trial at 50 dah was quite similar to trial B and larger than in trial A, regardless of the rearing system (Fig 9a and b) In trial C, the evolution of SGR followed a tendency similar to that in a previous trial, showing a hyperbolic curve evolution during the entire study while a decreasing trend was observed in the semi-intensive one regardless of the weaning protocol Finally, average SGR were no signi¢cantly di¡erent between rearing systems or weaning protocols (Fig 10a and b) r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 Aquaculture Research, 2010, 41, 433^449 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al SMIS-PLD IS-PLD 125 Prey per larvae (P LD); Prey/larvae/day Larval density (LD); (Larvae/litter) 150 100 75 50 SMI-LD SI-LD 25 20 15 35 Age (dah) Figure Estimated larval density evolution and available live prey (rotifers1Artemia) per day according to the rearing system (a) 35 Total length (mm) Survival was higher in semi-intensive system larvae regardless of the weaning protocol (Fig 11a and b) However, in weaning protocol (W2) survival improved signi¢cantly, particularly in the intensive system (12.5% inW2) ISW1-TC SMISW1-TC 30 25 20 15 Activity test for trial C 10 0 10 20 30 40 Age (dah) 50 60 30 Age (dah) 50 60 (b) 35 ISW2-TC SMISW2-TC Total length (mm) 30 25 20 15 10 At 20 dah, regardless of the weaning protocol applied, larvae reared under semi-intensive system conditions showed higher survival after the activity test than larvae reared in the intensive system Regarding the e¡ect of the weaning protocol, larvae fed according to the weaning protocol showed a higher survival after the activity test than those fed protocol for both rearing systems (Fig 12) At 30 dah, in larvae fed according to weaning protocol 1, no signi¢cant di¡erences were observed in survival among rearing systems However, in larvae fed following weaning protocol 2, survival of the larvae from the semi-intensive system was signi¢cantly higher than larvae from the intensive system (Fig 13) 0 10 20 40 Figure (a, b) Total length evolution (TL) of red porgy (Pagrus pagrus) larvae culture under di¡erent rearing systems and weaning protocols in trial C Ãsigni¢cant di¡erences Larval growth ¢ts to an exponential equation TL a  expbd, whereTL is the total length (mm), d is the age (days after hatching) and a, b were constants (Table 4) FA composition of P pagrus larvae Trial A In Trial A, the lipid contents of intensive systemreared larvae were signi¢cantly higher than the semi-intensive ones at 12 and 20 dah The n-3, n-3 r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 439 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al (a) 120 (a) 10 SMISW1-TC ISW1-TC SMISW1-TC ISW1-TC 100 % Survival SGR 80 60 40 * * * 20 0 5-15 Hatching 15-30 30-50 Total (5-50) Age (range) 20 35 Age (dah) 50 (b) 120 (b) 10 SMISW2-TC ISW2-TC SMISW2-TC ISW2-TC 100 % Survival SGR Aquaculture Research, 2010, 41, 433^449 80 * 60 * 40 * 20 0 Hatching 15-30 30-50 Total (5-50) Age (range) Figure 10 (a, b) Speci¢c growth rate evolution according to the rearing system and weaning protocol in trial C HUFA and DHA levels of red porgy larvae were significantly higher in the semi-intensive than in the intensive system on these days (12^20 dah), whereas no signi¢cant variation was observed at older stages (35 and 50 dah) (Table 5) Consequently, DHA/ARA, Oleic/DHA, Oleic/n-3 HUFA and n-3/n-6 ratios were signi¢cantly a¡ected at 12 dah At 50 dah, the 18:2n-6 level was signi¢cantly lower and the 18:3n-3 level was signi¢cantly higher in semi-intensive-reared larvae in comparison with intensivereared ones 20 35 Age (dah) 50 Figure 11 (a, b) Survival evolution according to the rearing system and weaning protocol in trial C 100 80 % Survival 5-15 60 40 20 Weaning Protocol Trial B In Trial B, at12 dah, the lipid content of intensive system-reared larvae was signi¢cantly higher than in semi-intensive ones On this day, the larval lipid level was higher than the level observed in previous trial A for both rearing systems As in the previous trial, the levels of n-3, n-3 HUFA and DHA of red porgy larvae were higher in the semi-intensive than in the intensive system at 12 and 20 dah, but not statistically 440 Figure 12 Survival 24 h after an activity test (10 s air exposure) of 20-day-old red porgy larvae according to the rearing system and weaning protocol in trial C signi¢cant The FA pro¢le of the larvae was not significantly a¡ected by the rearing systems at older stages (Table 6) As occurred in trial A, at 50 dah, the level of 18:2n-6 was signi¢cantly higher in the intensive r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 Aquaculture Research, 2010, 41, 433^449 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al 120 Discussion * % Survival 100 80 * 60 40 20 Weaning Protocol Figure 13 Survival 24 h after an activity test (1min air exposure) of 30-day-old red porgy larvae, according to the rearing system and weaning protocol in trial C Ãsigni¢cant di¡erences among rearing systems system-reared larvae with respect to semi-intensive system ones Trial C In Trial C, at 20 dah, the lipid content of intensive system-reared larvae was higher than in semi-intensive ones, although only signi¢cantly in the protocol fed larvae As in the previous trial, the levels of n-3 were higher in the semi-intensive than in the intensive system at 20 dah regardless of the weaning protocol No e¡ects were observed in n-3 HUFA and other individual FA at 20 dah when weaning protocol was used (Table 7); however, some di¡erences in the proportions of total saturates and monounsaturated FA among systems were found when weaning protocol was applied (Table 8) Di¡erences were mainly due to 16:0 and 18:0, which were 16.8% and 10.3% in the semi-intensive larvae and 19.4% and 11.3% in the intensive-reared larvae Besides, a signi¢cantly higher level of 18:3n-3 was observed in semi-intensive-reared larvae; moreover, a generally lower level of 18:2n-6 was observed in this system in comparison with intensive-reared larvae when weaning protocol was applied In addition, a lower content of 18:3n-3 and a higher 18:2n-6 level were associated with the early microdiet introduction for both rearing systems In relation to EFA, the level of DHA (19.8%) was signi¢cantly increased in larvae from the intensive system at 20 dah when protocol was used; moreover, this increase was evident in comparison with larvae fed on weaning protocol and cultured on the same rearing system Major FAs in the whole body of red porgy larvae were 22:6n-3, 16:0, 18:1n-9, 18:0, 18:2n-6, 20:5n-3, 16:1n-7, 18:1n-7, 18:3n-3 and 20:4n-6 Similar results have been reported for other marine species such as gilthead seabream (Roo, HernaŁndez-Cruz, FernaŁndezPalacios & Izquierdo 2005), barramundi, Latris lineata F., (Bransden, Battaglene, Morehead, Dunstan & Nichols 2005) or Atlantic cod (Plante, Pernet, Hache¤, Ritchie, Ji & McIntosh 2007) The importance of longchain FAs such as DHA (22:6n-3), EPA (20:5n-3) and ARA (20:4n-6) had been widely studied in marine ¢sh species (Izquierdo 1996; Sargent et al 1997), playing an important role in the cell membrane structure, function and development of di¡erent tissues and organs as neural and visual systems at early stages The proportions of DHA in red porgy larvae at 12 dah were almost two times higher than those observed in the live preys Thus, the level of DHA in larval tissues (15^20%) in early stages (12^20 dah) largely exceeded the values found in the diet (9^10%) (Table 2) These results agreed with the observations of Izquierdo (1996), suggesting a selective incorporation of this FA into both lipid reserves and membranes In contrast, ARA remains almost constant and EPA clearly re£ects dietary changes during ontogeny The proportion of EPA in red porgy feeding on Artemia and microdiet increased signi¢cantly in comparison with the values recorded for early stages due to a higher EPA content in this type of food Consequently, the DHA/EPA decreased with age and the EPA/ARA ratio tended to increase in older larvae Regardless, of the rearing system and trial, three di¡erent periods could be found along larval development: a ¢rst one (5^15 dah) with high growth rates (6%) despite being a very sensitive stage for larval survival; a second period (15^30 dah), related to completion of metamorphosis, with lower growth rates (2, 3% for trial A and around 4%, for B and C) and also quite sensitive to mortalities; and, ¢nally, a third post-metamorphic one (30^50 dph) with increased growth rates (5%) and low mortality During the ¢rst period (5^15 dah), larval performance seemed to be closely related to the success after the onset of exogenous feeding and the yolk reserves’ depletion Thus, the low growth and survival ratios obtained in semi-intensive larvae in trial A could be due to an insu⁄cient prey intake, which was also re£ected in the larval FA composition Indeed, retention of DHA, together with reduction in 18:0 and 18:1n-9, has been found to be associated r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 441 442 0.38 0.49 0.40a 0.03a 0.01a 0.16a 2.92 2.90 8.45 0.77 0.51 2.30 30.58 26.14 28.89 12.59 14.95 24.23 Æ Æ Æ Æ Æ Æ 0.38 0.10 0.81b 0.01b 0.03b 0.05b 0.35 1.01 0.96 0.14 1.17 1.12 2.15 2.72 5.85 0.75 0.50 2.60 30.49 29.18 26.93 10.39 13.92 24.18 17.51 5.01 9.78 12.08 7.94 5.90 2.19 1.19 2.74 5.90 16.04 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.00 0.04 0.07 0.01 0.00 0.11 0.85 0.97 0.25 0.53 0.55 0.34 0.50 0.09 0.21a 0.24 0.47 0.09 0.04 0.41 0.04 0.09 0.21a 2.36 1.84 4.33 0.78 0.44 2.63 29.87 27.68 29.55 11.25 13.74 27.91 17.05 6.15 10.17 12.34 7.37 5.48 1.48 1.14 3.68 8.66 15.91 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.02 0.14 0.38 0.07 0.03 0.11 0.13 0.84 1.05 0.09 0.39 1.01 0.33 0.24 0.21b 0.40 0.17 0.23 0.03 0.01 0.10 0.16 1.01b 17.22 Æ 2.10 76.73 Æ 5.06 2.21 Æ 0.46 SI-TA 3.10 2.12 6.54 0.82 0.51 2.05 33.97 21.28 28.82 14.06 12.62 21.62 21.16 2.27 8.66 10.97 5.45 10.04 5.18 1.46 2.05 6.36 13.41 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.17 0.25 0.41 0.04 0.01 0.04 1.22 0.20 0.12 0.29 0.09 0.28 0.99 0.09 0.09 0.25 0.27 0.23 0.06 0.10 0.07 0.55 0.41 15.07 Æ 0.85 74.03 Æ 7.82 2.75 Æ 0.29a SMI-TA 35 dah 3.25 1.19 3.89 0.95 0.45 2.46 28.68 23.05 31.97 12.98 12.64 22.47 18.61 3.71 7.86 10.00 4.58 8.22 8.26 1.71 2.72 8.84 10.55 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.05 0.01 0.09 0.03 0.02 0.05 0.02 0.04 0.05 0.25 0.09 0.51 0.07 0.26 0.02 0.14 0.37 0.12 0.43 0.01 0.11 0.24 0.19 14.50 Æ 0.60 81.23 Æ 5.29 3.35 Æ 0.21a SI-TA 5.70 1.50 8.52 1.28 0.69 3.66 27.83 30.57 31.92 8.75 17.31 21.74 16.87 5.07 7.56 15.05 6.69 6.51 8.49 2.15 1.41 7.88 11.79 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.95 0.01 1.37 0.01 0.03 0.34 0.13 1.37 1.79 0.33 0.22 0.91 0.49 0.43 0.20 0.07 0.63a 0.16 0.76a 0.24 0.21 0.14 0.14 19.21 Æ 0.57a 64.59 Æ 4.77 3.43 Æ 0.48 SMI-TA 50 dah Values (mean Æ SD) followed by di¡erent superscript letters within a row for the same trial were signi¢cantly di¡erent (Po0.05) DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid; ARA, arachydonic acid; HUFA, highly unsaturated fatty acid; SMIS, Semi-intensive system; IS, Intensive system; TA, Trial A 3.08 2.66 8.08 0.66 0.43 2.72 Æ Æ Æ Æ Æ Æ EPA/ARA DHA/EPA DHA/ARA Oleic/DHA Oleic/n-3HUFA n-3/n-6 1.43 1.83 0.31 0.68 0.31 0.84 Æ Æ Æ Æ Æ Æ 28.28 25.37 32.20 11.85 13.53 27.70 SSaturated SMono-unsaturated Sn-3 Sn-6 Sn-9 Sn-3HUFA Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 1.63 0.32 0.58a 0.12 0.85a 0.91 0.24 0.08 0.04 0.93 0.62a Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 15.19 5.79 9.88 11.81 5.06 7.68 3.81 1.28 2.21 6.83 17.87 16:00 16:1n-7 18:00 18:1n-9 18:1n-7 18:2n-6 18:3n-3 20:1n-9 ARA (20:4n-6) EPA (20:5n-3) DHA (22:6n-3) 0.64 0.13 0.86b 0.31 0.38b 0.15 0.11 0.31 0.19 0.14 0.18b 18.40 Æ 0.52b 78.77 Æ 3.00 1.65 Æ 0.34 14.67 Æ 0.27a 83.27 Æ 4.07 1.64 Æ 0.32 % Lipids (dw) % Protein (dw) % Ash (dw) 17.58 6.54 10.02 12.44 3.95 9.08 4.00 1.53 1.92 5.56 16.13 SMI-TA SI-TA SMI-TA 16.77 Æ 0.12 81.07 Æ 4.94 2.07 Æ 0.23 20 dah 12 dah Days after hatching Treatment 6.04 1.47 8.85 1.28 0.64 3.53 28.61 28.07 32.99 9.34 18.27 24.24 17.04 5.39 7.93 15.57 4.22 7.13 6.67 2.58 1.38 8.32 12.20 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.00 0.00 0.00 0.09 0.06 0.10 1.47 0.57 1.21 0.25 0.64 1.10 0.71 0.41 0.87 0.73 1.18b 0.24 0.22b 0.09 0.05 0.28 0.40 17.81 Æ 0.23b 68.80 Æ 2.57 3.79 Æ 0.47 SI-TA Table Proximate composition [lipid, protein and ash content; g kg À dry weight (dw)] and selected fatty acid composition (% total fatty acids) of whole-body red porgy larvae at 12, 20, 35 and 50 dah in trial A Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al Aquaculture Research, 2010, 41, 433^449 r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 2.53 3.56 8.97 0.69 0.48 2.15 0.09a 0.04 0.24a 0.03a 0.02 0.07 Æ Æ Æ Æ Æ Æ 18.12 4.98 9.87 11.25 4.24 8.57 3.47 1.18 1.82 4.59 16.30 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.11b 0.15 0.02b 0.01b 0.00 0.01 1.65 0.95 0.28 0.20 0.29 0.13 1.26 0.47 0.36 0.09b 0.22 0.16b 0.34 0.07 0.03 0.13 0.25 2.09 3.61 7.53 0.77 0.54 2.60 33.10 23.45 27.48 10.55 15.41 24.63 20.14 3.98 10.86 13.27 4.28 6.16 2.27 0.95 2.30 4.82 17.27 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.12 0.34a 0.28a 0.16a 0.13 0.02a 1.89 1.85 2.49 0.99 2.01 1.72 1.94 0.01 0.12 2.16a 0.16 0.74 0.69 0.11 0.18 0.63 0.68 2.13 3.66 7.80 0.73 0.52 2.55 35.30 23.07 26.41 10.38 13.84 22.51 20.87 4.03 12.21 11.70 4.77 6.14 3.25 0.97 2.05 4.37 15.98 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.00 0.07b 0.15b 0.05b 0.03 0.10b 0.60 0.13 0.12 0.40 0.55 0.16 0.88 0.42 0.25 0.69b 0.13 0.35 0.21 0.04 0.02 0.05 0.15 14.68 Æ 0.63 79.27 Æ 2.62 2.12 Æ 0.10 IS-TB 2.95 2.21 6.52 1.25 0.79 1.58 32.18 25.92 24.02 15.27 16.74 18.33 20.54 2.69 9.31 14.37 4.78 12.34 4.80 1.54 1.77 5.23 11.52 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.07 0.07 0.36 0.06 0.05 0.15 0.13 0.38 1.14 0.78 0.45 0.87 0.46 0.01 0.37 0.23 0.31 1.06 0.25 0.01 0.16 0.34 0.41 15.71 Æ 0.30 69.29 Æ 2.79 3.23 Æ 0.11a SMI-TB 35 dah 2.66 2.58 6.74 1.18 0.77 1.81 33.05 25.96 24.48 13.92 16.97 19.30 20.96 2.93 9.81 14.76 4.62 10.62 4.43 1.65 1.91 4.94 12.74 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.51 0.45 0.62 0.19 0.10 0.46 1.04 0.89 1.97 2.43 0.54 2.59 0.43 0.30 1.26 0.30 0.39 3.06 0.88 0.26 0.43 0.31 2.26 17.13 Æ 1.32 67.25 Æ 4.50 3.48 Æ 0.22b IS-TB 5.29 1.60 8.53 1.34 0.73 1.66 32.00 28.56 22.80 13.84 17.50 20.05 20.60 5.23 7.59 14.65 3.17 11.24 2.36 2.27 1.32 6.86 10.95 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.60 0.12 1.62 0.04 0.01 0.20 0.70 0.28a 1.39 0.81 0.37 1.34 0.09 0.04 0.25 0.71 0.48 1.05a 0.03 0.22a 0.27 0.65 0.19 17.64 Æ 1.10 63.12 Æ 0.85 3.70 Æ 0.22 SMI-TB 50 dah Values (mean Æ SD) followed by di¡erent superscript letters within a row for the same trial were signi¢cantly di¡erent (Po0.05) DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid; ARA, arachydonic acid; HUFA, highly unsaturated fatty acid; SMIS, Semi-intensive system; IS, Intensive system; TB, Trial B 2.56 3.70 9.48 0.76 0.53 2.41 1.38 0.84 0.63 0.11 0.02 0.62 Æ Æ Æ Æ Æ Æ 29.56 23.35 28.14 11.68 15.63 24.36 SSaturated SMono-unsaturated Sn-3 Sn-6 Sn-9 Sn-3HUFA EPA/ARA DHA/EPA DHA/ARA Oleic/DHA Oleic/n-3HUFA n-3/n-6 29.40 23.16 27.38 12.73 14.42 23.37 1.10 0.53 0.30 0.18a 0.20 0.10a 0.10 0.05 0.01 0.17 0.47 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 19.47 4.55 8.59 12.81 3.61 7.79 3.16 1.12 1.79 4.58 16.92 16.20 Æ 1.35 87.43 Æ 9.69 2.27 Æ 0.13 16:0 16:1n-7 18:0 18:1n-9 18:1n-7 18:2n-6 18:3n-3 20:1n-9 ARA (20:4n-6) EPA (20:5n-3) DHA (22:6n-3) 20.45 Æ 1.03 71.10 Æ 3.46a 1.64 Æ 0.18 17.37 Æ 0.91 85.95 Æ 5.94a 1.69 Æ 0.25 % Lipids (dw) % Protein (dw) % Ash (dw) a SMI-TB IS-TB SMI-TB a 20 dah 12 dah Days after hatching Treatment 3.66 2.26 8.16 1.22 0.78 1.56 30.62 27.07 24.40 15.74 17.23 18.94 19.88 3.14 8.42 14.59 4.37 12.57 4.42 2.11 1.53 5.37 12.04 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 1.23 0.15 2.31 0.13 0.09 0.20 2.81 0.61b 1.86 1.18 0.28 1.99 2.12 0.65 1.17 0.46 0.63 1.64b 0.90 0.64b 0.32 0.81 1.06 16.07 Æ 0.57 63.89 Æ 2.05 3.43 Æ 0.14 IS-TB Table Proximate composition [lipid, protein and ash content; g kg À dry weight (dw)] and selected fatty acid composition (% total fatty acids) of whole-body red porgy larvae at12, 20,35 and 50 dah in trial B Aquaculture Research, 2010, 41, 433^449 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al 443 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al Aquaculture Research, 2010, 41, 433^449 Table Proximate composition [lipid, g kg À dry weight (dw)] and selected fatty acid composition (% total fatty acids) of whole-body red porgy larvae at 20, 35 and 50 dah in trial C according to the weaning protocol 20 dah 35 dah Days after hatching Treatment SMIW1-TC ISW1-TC 50 dah SMIW1-TC 17.42 Æ 1.35 18.73 Æ 1.46 SMIW1-TC ISW1-TC 19.63 Æ 5.99 17.03 Æ 0.44 % Lipids (dw) 15.21 Æ 1.25 16:0 16:1n-7 18:0 18:1n-9 18:1n-7 18:2n-6 18:3n-3 20:1n-9 ARA (20:4n-6) EPA (20:5n-3) DHA (22:6n-3) 17.06 4.14 9.88 13.33 5.04 6.16 4.19 1.19 2.64 6.24 15.25 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.52 0.25 0.27 0.27 0.22 0.21 0.20 0.09 0.10 0.16 0.80 17.22 3.89 10.21 12.79 4.35 7.99 2.06 1.35 2.20 4.87 17.31 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.35 0.02 0.13 0.09 0.00 0.00 0.03 0.01 0.00 0.02 0.22 15.90 3.37 8.05 15.72 4.74 7.42 6.89 1.55 2.08 6.41 14.28 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 1.55 0.14a 0.72a 0.52 0.25 0.52 0.64 0.09a 0.07 0.10 0.21 18.95 2.35 10.82 15.86 4.51 6.52 8.64 1.09 1.62 4.97 10.68 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 5.15 0.44b 1.48b 0.98 0.71 1.06 1.34 0.18b 0.30 0.95 1.79 19.30 5.34 6.88 11.98 3.61 10.49 2.84 1.83 1.22 8.33 13.60 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.81a 0.17 0.23 0.40a 0.13 0.30 0.15 0.01 0.03 0.05 0.28 20.55 5.43 7.99 14.10 3.34 11.53 1.98 2.13 1.34 7.21 12.38 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.16a 0.32 0.72 1.24a 0.56 1.16 0.66 0.34 0.27 0.87 2.44 SSaturated SMono-unsaturated Sn-3 Sn-6 Sn-9 Sn-3HUFA 29.27 26.49 29.98 11.05 15.46 24.08 Æ Æ Æ Æ Æ Æ 0.70 0.34 0.14 0.48 0.25 0.06 30.86 25.50 27.83 11.94 15.98 24.35 Æ Æ Æ Æ Æ Æ 0.38 0.05 0.22 0.05 0.16 0.24 26.75 27.70 31.47 11.13 18.18 22.75 Æ Æ Æ Æ Æ Æ 2.28 0.95 1.06 0.48 0.61 0.34 32.74 26.30 28.44 9.74 18.04 17.55 Æ Æ Æ Æ Æ Æ 7.14 1.46 4.53 1.43 0.94 3.08 30.50 25.40 29.10 13.12 14.02 23.99 Æ Æ Æ Æ Æ Æ 1.11 0.79 0.36a 0.04 0.44 0.44 32.08 27.46 23.99 14.22 16.54 21.54 Æ Æ Æ Æ Æ Æ 0.72 1.88 2.43a 1.04 1.60 2.83 2.36 2.45 5.78 0.87 0.55 2.72 Æ Æ Æ Æ Æ Æ 0.05 0.07 0.21 0.02 0.01 0.10 2.22 3.55 7.87 0.74 0.53 2.33 Æ Æ Æ Æ Æ Æ 0.01 0.06 0.09 0.00 0.00 0.01 3.09 2.23 6.87 1.10 0.69 2.83 Æ Æ Æ Æ Æ Æ 0.10 0.03 0.14 0.03 0.02 0.12 3.07 2.15 6.61 1.52 0.93 2.92 Æ Æ Æ Æ Æ Æ 0.06 0.07 0.15 0.35 0.22 0.17 6.82 1.63 11.13 0.88 0.50 2.22 Æ Æ Æ Æ Æ Æ 0.10 0.02 0.00 0.01 0.01 0.03a 5.46 1.71 9.44 1.18 0.66 1.69 Æ Æ Æ Æ Æ Æ 0.64 0.22 2.15 0.28 0.12 0.20b EPA/ARA DHA/EPA DHA/ARA Oleic/DHA Oleic/n-3HUFA n-3/n-6 16.58 Æ 3.30 ISW1-TC Values (mean Æ SD) followed by di¡erent superscript letters within a row for the same trial were signi¢cantly di¡erent (Po0.05) DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid; ARA, arachydonic acid; HUFA, highly unsaturated fatty acid; SMIS, Semiintensive system; IS, Intensive system; TC Trial C with starvation in marine ¢sh larvae (Koven, Kissil & Tandler 1989; Izquierdo 1988; Rodriguez 1994) Accordingly, the increase in prey density during this period in the semi-intensive system in trial B (4^ rot mL À 1) in comparison with trial A (1^ rot mL À 1) markedly decreased mortalities and improved growth rates Moreover, on the one hand, wild zooplankton aggregations are usually over ind mL À (Browman 2005) and on the other, the larval visual perception ¢eld (VPF; water volume where the larvae is able to visually detect a prey) (Gallager, Yamazaki & Davis 2004) for a marine ¢sh larvae of this size (5 mm) is not 41mL (Browman & Skiftesvik 1996; Galbraith, Browman, Racca, Skiftesvik & StPierre 2004) Therefore, prey densities under rot mL À in a semi-intensive system in trial A would be too low to promote predator^prey encounters Accordingly, in Trial B, rotifer density was increased over rot mL À to satiate the feed intake of red porgy, which, in addition, seems to be more voracious than other sparids (HernaŁndez-Cruz et al 444 1999) However, the low growth and high mortalities recorded during this period in the intensive system could be related more to other causes, such as unsuccessful use of prey nutrients under more stressful rearing conditions or related to changes in the bacterial community structures in the rearing water as was recently reported by Nakase, Nakagawa, Miyashita, Nasu, Senoo, Matsubara and Eguchi (2007) These authors found that poor growth and early mortality (4^14 dah) in a close species to red porgy such as Japanese red seabream, Pagrus major, reared under an intensive system was associated with the increase in g-Proteobacteria in seawater despite the daily addition of Nannochloropsis sp., and suggested a marked e¡ect of the microalgae physiological condition on the bacterial community structures In the present study, Nannochloropsis sp was added daily in both systems, but the concentration and physiological condition of this microalga were markedly affected by the speci¢c characteristics of each system (tank volume and depth, water turnover and turbu- r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 Aquaculture Research, 2010, 41, 433^449 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al Table Proximate composition [lipid, g kg À dry weight (dw)] and selected fatty acid composition (% total fatty acids) of whole-body red porgy larvae at 20, 35 and 50 dah in trial C according to the weaning protocol 20 dah 35 dah 50 dah Days after hatching Treatment SMIW2-TC ISW2-TC SMIW2-TC ISW2-TC SMIW2-TC ISW2-TC % Lipids (dw) 15.87 Æ 2.01a 18.00 Æ 1.47b 15.33 Æ 5.29 17.23 Æ 2.42 14.84 Æ 2.02 16.00 Æ 2.11 16:0 16:1n-7 18:0 18:1n-9 18:1n-7 18:2n-6 18:3n-3 20:1n-9 ARA (20:4n-6) EPA (20:5n-3) DHA (22:6n-3) 16.81 3.99 10.13 12.39 4.60 7.74 2.69 1.19 2.41 5.59 16.81 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.14a 0.16 0.35a 0.42 0.09a 0.40 0.15a 0.08 0.04 0.06 0.41a 19.49 3.54 11.34 12.05 3.84 7.42 1.21 1.15 2.24 4.40 19.08 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.83b 0.08 0.37b 0.05 0.12b 0.78 0.16b 0.19 0.06 0.10 0.67b 17.59 3.17 8.56 13.67 3.96 8.84 4.11 1.47 2.21 5.95 17.38 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.85a 0.09 0.33 0.28 0.18 0.53 0.48a 0.02 0.10 0.13 0.25 19.09 2.56 8.91 11.84 3.38 14.48 2.38 1.64 1.78 5.39 17.71 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.40b 0.13 1.29 0.22 0.16 2.98 0.54b 0.35 0.33 0.51 1.32 18.82 5.43 6.89 11.81 3.43 11.07 2.00 2.02 1.17 8.61 13.97 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 0.33 0.28 0.24 0.41 0.14 0.30 0.13a 0.04 0.03 0.35a 0.42a 20.33 5.26 7.56 12.15 3.20 11.30 1.28 2.14 1.20 7.84 14.94 Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ Æ 1.28 0.03 0.62 0.73 0.08 0.18 0.02b 0.57 0.02 0.13b 0.22b SSaturated SMono-unsaturated Sn-3 Sn-6 Sn-9 Sn-3HUFA 29.99 25.34 28.98 12.19 14.70 24.85 Æ Æ Æ Æ Æ Æ 0.43a 0.62a 0.43 0.32 0.36 0.52 34.40 23.43 28.07 11.45 14.14 25.45 Æ Æ Æ Æ Æ Æ 1.35b 0.52b 0.87 0.96 0.24 0.93 29.18 24.50 31.00 12.58 16.08 25.30 Æ Æ Æ Æ Æ Æ 1.47 0.48 0.88a 0.51a 0.31a 0.36 30.57 21.29 28.50 17.35 14.19 24.84 Æ Æ Æ Æ Æ Æ 1.87 0.16 0.47b 2.50b 0.30b 0.93 30.31 25.39 28.86 13.32 13.93 24.68 Æ Æ Æ Æ Æ Æ 0.70 0.81 0.13a 0.22 0.35 0.10 32.02 25.34 27.61 13.37 14.29 24.69 Æ Æ Æ Æ Æ Æ 2.10 1.98 0.29b 0.18 1.30 0.32 2.32 3.00 6.98 0.74 0.50 2.38 Æ Æ Æ Æ Æ Æ 0.03a 0.06a 0.07a 0.04a 0.03 0.10 1.96 4.34 8.53 0.63 0.47 2.47 Æ Æ Æ Æ Æ Æ 0.01b 0.10b 0.19b 0.02b 0.02 0.27 2.69 2.92 7.87 0.79 0.54 2.47 Æ Æ Æ Æ Æ Æ 0.08 0.06 0.29a 0.01 0.00 0.12a 3.13 3.32 10.10 0.67 0.48 1.67 Æ Æ Æ Æ Æ Æ 0.78 0.58 1.02b 0.06 0.03 0.29b 7.39 1.63 11.97 0.85 0.48 2.17 Æ Æ Æ Æ Æ Æ 0.46a 0.12a 0.15a 0.05 0.02 0.04a 6.51 1.90 12.40 0.81 0.49 2.06 Æ Æ Æ Æ Æ Æ 0.12b 0.03b 0.02b 0.05 0.03 0.04b EPA/ARA DHA/EPA DHA/ARA Oleic/DHA Oleic/n-3HUFA n-3/n-6 Values (mean Æ SD) followed by di¡erent superscript letters within a row for the same trial were signi¢cantly di¡erent (Po0.05) DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid; ARA, arachydonic acid; HUFA, highly unsaturated fatty acid; SMIS, Semiintensive system; IS, Intensive system; TC, Trial C lence and light distribution) and could a¡ect the bacterial community structures to a less favourable one in the intensive system In the second period (15^ 30 dah), morphological and physiological changes led to an improvement in many biological functions such as digestion and vision Thus, during this period extracellular digestion improves, gastric glands start to become functional (Govoni, Boelhert & Watanabe 1986; Segner, Storch, Reinecke, Kloas & Hanke 1994; Roo, Socorro, Izquierdo, Caballero, FernaŁndez & FernaŁndez-Palacios1999; Socorro, Izquierdo, FernaŁndez, FernaŁndez-Palacios, Caballero & Roo 1999; Darias, Murray, Mart|¤ nez-Rodr|¤ guez, CaŁrdenas & Yu¤fera 2005) and a new type of photoreceptor (rods) develops, increasing retina sensitivity necessary for wild ¢sh to shift from shallow to deep waters (Roo et al 1999) At the same time, important changes in the rearing techniques occur in feeding (switch in live prey, from rotifer toArtemia and weaning to dry feed) and water quality (green to clear water) Very low growth rates (2%) were found during this period in both rearing systems in trial A, which were increased in trial B (4%) when rotifer feeding was prolonged up to 30 dah (20 dah in trial A) and the 24-h photoperiod was reduced to 12 h from 20 dah (40 dah in trial A) Inadequate illumination levels with continuous photoperiods and the progressive change to clear water in such a sensitive phase could act as a stressor for the larval population (Boeuf & Le Bail 1999) But the better growth rate found in trial B could also be related to the better nutritional value of rotifers in comparison with Artemia Indeed, red porgy ability to digest Artemia was very de¢cient in both trials A and B, as was observed under a binocular microscope (Fig.1) The poor digestion of Artemia could be related to the poor development of red porgy gut at this stage, where, despite having a very rudimentary stomach and gastric glands as early as 19 dah (Roo et al 1999; Socorro et al 1999; Socorro 2006), acid digestion is not completely functional until 35 dah (Darias, Murray, Gallant, Susan, Douglas, Yu¤fera & Mart|¤ nez-Rodr|¤ guez 2007) In addition, this reduced Artemia r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 445 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al digestion could also be related to an excessive intake of this prey by the voracious red porgy larvae, which would reduce the intestinal transit time of Artemia in the larval gut In agreement with the observed reduction in prey digestion when larvae were fed Artemia, the red porgy DHA content remained constant during rotifer feeding (12^20 dah) and was markedly reduced with Artemia feeding (35^50 dah), although a similar content of this FA was recorded in both preys (Table 2) Previous studies have shown that red porgy has a high DHA requirement during larval development, its reduction in live preys causing severe skeletal anomalies (Roo et al 2009) Moreover, in weaning protocol 2, co-feeding with a dry diet occurred as early as 15 dah and this could also be related to the improvement in survival in this second period, indicating the superior nutritional quality of the commercial diet over Artemia at this larval stage In fact, the use of an early co-feeding protocol signi¢cantly a¡ected larval FA composition, increasing the level of DHA in early co-fed larvae reared under an intensive system with respect to control larvae The bene¢ts of co-feeding protocols were reported previously in other species such as gilthead seabream (Kolkovski, Koven & Tandler 1997; Rosenlund, Stoss & Talbot 1997; Koven, Kolkovski, Hadas, Gamsiz & Tandler 2001), or barramundi, Lates calcacifer B (Curnow, King, Bosmans & Kolkovski 2006) In this study, the early microdiet introduction was found to be more effective in the intensive system than in the semi-intensive one, and was also re£ected in the larval FA composition, which was higher in 18:2n-6 in early weaned larvae After complete metamorphosis and weaning to a dry diet, third period (30^50 dph), growth rates were good in all trials and similar to the second stage These results seemed to be related to the higher maturation of the ¢sh, particularly of the digestive, neural, sensorial and endocrine systems (Roo et al 1999; Socorro et al 1999; Socorro 2006; Darias et al 2005, 2007) Overall, growth rates until older stages allowed attaining a body weight at 95 dah of 6.2 Æ 1.5 g, values signi¢cantly higher than in Sparus aurata juveniles reared under similar conditions (F.J Roo, C.M HernaŁndez-Cruz, J.A Socorro, H FernaŁndez-Palacios & M.S Izquierdo, unpubl obs.), and in agreement with the results of Papandroulakis, Kentouri and Divanach (2004) During this third period, the main factor threatening survival seemed to be the apparition of cannibalistic behaviour, which, in this study, was observed at 8^8.5 mm total length 446 Aquaculture Research, 2010, 41, 433^449 although not causing signi¢cant losses Cannibalistic behaviour also appears in other sparids such as common dentex, Dentex dentex L (Koumoundouros, Divanach & Kentouri 1999), or gilthead seabream Although in the latter cannibalism is reduced by grading and separation of di¡erent ¢sh sizes, manipulation at this stage causes sudden mortalities in several species such as common dentex (Mourente, Tocher, Diaz-Salvago, Grau & Pastor 1999) In this sense, sudden mortalities by the shock syndrome were also found in red porgy when an acute stress (activity test) was applied, but being lower in ¢sh reared under the semi-intensive system The apparition of the shock syndrome after acute stress has been suggested to be a consequence of nutritional imbalances during previous larval stages (Izquierdo, Watanabe, Takeuchi, Arakawa & Kitajima 1989), in relation to essential FAs, particularly DHA, as it seems to be the case for red porgy (Roo et al 2009), which had a higher DHA content when reared in semi-intensive systems in agreement with the higher survival after an acute stress test In a wide range of marine species, the use of semiintensive technology for larval rearing results in higher survival and growth performance than in intensive larval rearing techniques (Papandroulakis, Kentouri, Maingot et al 2004; Roo et al 2005) Several factors have been held to be responsible for this marked di¡erence in larval performance between both systems such as the more stable rearing conditions in the semi-intensive one, which allows to develop more mature bacterial £ora bu¡ering the e¡ect of daily bacterial charges included with live prey addition Nutrition during the second period could also be an important factor related to the better performance of ¢sh in a semi-intensive system Thus, survival of intensive system red porgy was much closer to that of semi-intensive ones only when weaning was moved ahead and Artemia Instar II density was reduced The main di¡erence in the larval success between both systems for this species remains in the rotifers’ feeding period, when the density of larvae is reduced from 125 to about 17 larvae L À at 15 dah, and in the semi-intensive system from to larvae L À At this stage, one of the most important di¡erences observed was related to prey availability in the intensive system (around 425 rotifers larva À day À in the intensive system and 1700 rotifers larva À day À in the semi-intensive system), together with those related to water turnover, green water condition and tank management Hence, higher survival during the rotifer feeding period in a r 2009 The Authors Journal Compilation r 2009 Blackwell Publishing Ltd, Aquaculture Research, 41, 433^449 Aquaculture Research, 2010, 41, 433^449 Advances in rearing techniques of red porgy Pagrus pagrus F J Roo et al semi-intensive system could be related to a better utilization of prey nutrients under more favourable environmental conditions and hence lower energy demands This hypothesis is supported by the longer yolk and lipid reserve duration (Papandroulakis, Kentouri, Maingot et al 2004), and the higher DHA content in semi-intensive system-reared larvae, although a lower EFA content was recorded in the unconsumed rotifers in this system (Table 3) in the present study Moreover, the higher levels of 18:3n-3 in semi-intensive larvae in comparison with intensive ones indicated a higher intake of rotifers that were particularly rich in this FA In contrast, less favourable culture conditions such as increasing stocking density may induce lower food consumption and utilization as reported for juvenile gilthead seabream and white sea bream (Montero, Izquierdo,Tort, Robaina & Vergara 1999; Montero, Robaina, Socorro, Vergara, Tort & Izquierdo 2001; Papoutsoglou, Karakatsouli, Pizzonia, Dalla, Polissidis & PapadopoulouDaifoti 2006) and higher energy demands, which could lead to higher nutritional requirements Conclusions The results of this study allowed improvement in the rearing protocols for red porgy, increasing the ¢nal total length and survival at 50 dah from the initial trial to the last ones from 18.9 to 25.13 mm and from 4.9% to 12.5% in the intensive system and from 23.52 to 26.4 mm and 4.4% to 28.7% in the semi-intensive system At present, the best larval rearing protocol to sustain a regular and predictable red porgy ¢ngerling demand is the semi-intensive system technology Besides, early co-feeding and weaning, together with a reduction in stressors during rotifer feeding, is recommended in this species to improve larval performance under an intensive system Acknowledgments Funding was partially provided by the Spanish Ministries of Science and Education (AGL2003^09131) and the Ministry of Agriculture, Fisheries and Food (Jacumar: Promocio¤n del cultivo de nuevas especies de espaŁridos: Ensayos piloto y transferencia tecnolo¤gica) J Roo, thanks the ¢nancial support provided by the Spanish Ministry of Science and Education and European Social Funds through the programme‘Incorporacio¤n 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Rombaur G., Sorgeloos P & Verstraete W (2000) Probiotic bacteria as biological control agents in r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 345^355 Aquaculture Research, 2010, 41, 345^355 Enhancing natural defences in aquaculture species J W Sweetman et al aquaculture Microbiology and Molecular Biology Reviews 64, 655^671 Waldroup P.W., Oviedo-Rondon... with receptors of similar glycation, e.g blood cells and enterocytes Because various receptors di¡er in glyca- r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 333^344 Aquaculture Research, 2010, 41, 333^344 An update on antinutrients in aquaculture feeds — Krogdahl et al tion, their susceptibility to a particular lectin may differ Hendriks, van den... trypsin activity Results within each parameter marked with di¡erent letters were signi¢cantly di¡erent (Po0.05) 338 r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 333^344 Aquaculture Research, 2010, 41, 333^344 An update on antinutrients in aquaculture feeds — Krogdahl et al Figure 6 The following molecular outline of a-galactosyl homologues of sucrose:... 2009) The work of Uran, Goncalves, Taverne-Thiele, Schrama,Verreth and Rombout (2008) indicates that r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 333^344 339 An update on antinutrients in aquaculture feeds — Krogdahl et al Aquaculture Research, 2010, 41, 333^344 Figure 7 Histological characteristics of distal intestinal mucosa from Atlantic salmon fed... dietary exposure to SBM More speci¢c investigations are needed to reveal which subtypes of T cells are present In any r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 333^344 Aquaculture Research, 2010, 41, 333^344 An update on antinutrients in aquaculture feeds — Krogdahl et al case a T-cell-mediated response appears to be involved in this example of a food-sensitive... intestinal and liver morphology in ¢ngerling rainbow trout Oncorhynchus mykiss Fisheries Science 74, 1075^ 1082 r 2010 The Authors Journal Compilation r 2010 Blackwell Publishing Ltd, Aquaculture Research, 41, 333^344 Aquaculture Research, 2010, 41, 333^344 An update on antinutrients in aquaculture feeds — Krogdahl et al Johnson I.T., Gee J.M., Price K., Curl C & Fenwick G.R (1986) In£uence of saponins... 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